Although the liver has long been considered a tolerogenic organ, most liver transplant recipients require lifelong immunosuppression, which can have negative consequences, such as recurrence of viral hepatitis or hepatocellular carcinoma. All transplant recipients, regardless of organ, have an increased risk of developing chronic kidney disease, infection, malignancies, or cardiovascular disease. Tolerance induction, therefore, would be a significant long-term benefit in terms of allograft and patient survival. We have successfully achieved renal allograft tolerance using a mixed chimerism approach in concert with a non-myeloablative regimen in both humans and non-human primates (NHPs). However, the clinical application of this approach in liver transplant recipients is not as practical or safe as in kidney transplant recipients because liver failure patients tend to be sicker at the time of transplantation and unable to withstand the toxicity of the conditioning regimen. However, if the bone marrow transplant is delayed and if we reduce the toxicity of the myelosuppressive conditioning regimen, these approaches can be safely applied. Cippa et al. recently reported a novel approach to hematopoietic stem cell (HSC) engraftment without myelosuppression using a B cell lymphoma-2 (Bcl-2) inhibitor. This novel approach has been successfully translated to an NHP kidney transplant model. Using an FDA-approved, highly selective Bcl-2 inhibitor, ABT-199 (venetoclax), with reduced dose (1.5 G) total body irradiation (TBI), we have been able to achieve significantly improved mixed chimerism and renal allograft tolerance in NHPs. Encouraged by these preliminary results, we hypothesize that mixed chimerism and liver allograft tolerance can be induced even more effectively without any myelosuppressive treatments. The primary objective of this proposal is to evaluate the efficacy of selective Bcl-2 inhibition to induce mixed chimerism for liver allograft tolerance in a clinically relevant NHP model. HLA mismatched cynomologus monkeys will be used as both donors and recipients for allogeneic orthotopic liver transplantation. All recipients will initially receive triple-drug immunosuppressive regimen (CyA, MMF, steroid) consistent with the peri-transplant period. Several months (2-3) after orthotopic liver transplant (OLTx), the recipients will undergo non-myeloablative conditioning (thymic irradiation, with or without ABT-199) and DBMT, followed by a short post-transplant course of Belatacept (20mg/kg on days 0, 2, 5 and 12) and one month course of CyA to see whether Bcl-2 inhibitor can induce mixed chimerism and liver allograft tolerance. We will then study the effect of Bcl-2 inhibition on alloreactive T lymphocytes and regulatory T cells to investigate the mechanism.